Download Chem*3560 Lecture 26: Cell adhesion and membrane fusion

Chem*3560
Lecture 26: Cell adhesion and membrane fusion
A distinguishing feature of complex organisms is the ability of cells to recognize each other and to adhere
together as multicellular tissues (Lehninger p.404). Several sets of integral membrane proteins help
mediate these interactions:
Integrins
Integrins are made up of one α-subunit and one β-subunit, anchored in the plasma membrane of
multicellular eukaryotes by one transmembrane helix each. The extracellular region forms a globular
domain which can bind and recognize the amino acid sequence -Arg-Gly-Asp- in the presence of Ca2+.
This characteristic sequence is found in a number of proteins that make up the extracellular matrix
(Lehninger p.313 Fig. 9-24). Extracellular matrix is the macromolecular network that holds cells
together in a tissue, and different tissues have different components of the extracellular matrix.
Extracellular matrix components
Proteoglycan - a combination of complex heteropolysaccharides such as chondroitin sulfate
and link proteins (Lehninger p.308-313). Also linked to cell membranes by
the protein syndecan (Lehninger p. 312).
Collagen
- a triple helical protein that forms the fibrous framework of bone and cartilege.
Fibronectin - fibrous proteins characteristic of the extracellular network of connective
Laminin
tissue (Lehninger p.313; the reference to laminin as a nuclear membrane
protein on p. 473 is incorrect. The nuclear proteins are Lamins).
There are 18 different α-integrins and 8 different β-integrins which are found paired up in various
specific combinations in particular cell and tissue types. These give each cell type its characteristic
affinities in different tissues.
When integrins bind to an appropriate component of
extracellular matrix, they communicate this to the
interior of the cell. This occurs in part because the αand β- integrin subunits realign with each other when
they bind their target, and partly because multiple integrins
cluster together at a site where binding can occur, and
this clustering of the intracellular domains is able to bring
together key regulatory protein tyrosine kinases, and to
serve as a point of anchorage for actin filaments of the
internal cytoskeleton (Lehninger p. 313, Fig. 9-24).
Cadherins are members of a family of Ca2+
binding proteins found on the plasma
membrane surface. The extracellular
structure consists of five consecutive
β-sheet domains with Asp-rich junctions
that bind Ca2+.
Ca2+ ions can serve as bridges between
two negative molecules, but β-sheets are
also designed to pair up so that a cadherin
only binds an identical cadherin on a
neighbouring cell. Cadherins bundle together
to give cells the ability to adhere to like cells
and make up a homogeneous tissue .
Ig-CAMs - Immunoglobulin-like cell
adhesion molecules are calcium
independent binding proteins. CAM
molecules have a transmembrane helix that is
connected to multiple immunoglobulin-like
domains on the extracellular surface. Immunoglobulin domains are antiparallel β-barrel structures that
perform protein-protein binding, more familiar as antibodies. N-CAM is neural cell specific, V-CAM
vascular cell adhesion molecule, etc.
Selectins - are extracellular adhesion molecules designed to bind to extracellular structural
polysaccharides, also with a Ca2+ dependent binding site, particlarly important in blood system.
Membrane fusion
Membrane fusion is an extension of adhesion processes, in which two membranes approach sufficiently
close to allow the bilayers to merge and the contents to combine.
Membrane fusion is involved in a variety of cellular processes that transfer contents from one cellular
compartment to another (Lehninger p.405):
Exocytosis: vesicles bud off from the Golgi apparatus and fuse with the plasma membrane to
allow secretion of proteins.
Endocytosis: vesicles bud off from the plasma membrane to fuse with endosome or lysosome
compartments.
Endosome-Lysosome fusion: merges endosomes with lysosome for final processing
Synaptic transmission: Synaptic vesicles (filled with neurotransmitter) fuse with synaptic
junction membrane to release neurotransmitter into synaptic cleft.
Viral infection: viral capsid fuses with plasma membrane
Sperm-egg fusion
Fusion of small vacuoles
Fusion occurs at specific target sites in the cell
Fusion specific proteins only occur at specific target locations, and the example here is the synaptic
junction. Fusion is mediated by membrane proteins that distort the approaching bilayers, which
otherwise repel each other due to negative charge.
The protein that opened up the discovery of the mechanism was recognized because of its sensitivity to
N-ethylmaleimide (NEM) which blocks enzymes, in particular ATPases, that depend on nucleophilic
acion of cysteine -SH.
NEM-Sensitive Fusion protein (NSF) is a cytoplasmic ATP-hydrolysing protein that is inactivated
by NEM, which then blocks the fusion process.
Soluble NSF attachment protein (SNAP) is a second cytoplasmic protein required for fusion.
SNAP receptors or SNAREs are helical transmembrane proteins that start the fusion process. Their
helical domains extend a long distance into the cytoplasm.
SNAREs come in two general types: v-SNAREs (vesicle-associated), are found in the vesicle
membrane. t-SNAREs (target associated) are found in the target region, e.g. inside the plasma
membrane, that will be fused to. Altogether, there are 30 types of SNAREs in mammalian cells, and
these would distinguish the many different vesicle/target systems in a cell (Lehninger p. 406).
When the vesicle approaches the target (it
may be brought there by components of the
cytoskeleton) v-SNAREs and t-SNAREs
associate by arranging the cytoplasmic
helices into a four-helix bundle, which
"docks" the vesicle into the target
membrane. (Only shown as two helices in
the schematic to keep figure simple; see the
detail structure below)
As the SNAREs wind up, they draw the
vesicle closer to the surface of the cell
membrane. Distortion of the bilayer by the
SNARE complex then causes bilayers to
merge, ultimately leading to fusion.
One v-SNARE is anchored in the vesicle
membrane by the transmembrane helix at its
C-terminal end. This binds to two
t-SNAREs that actually form three helices.
The single helix t-SNARE is anchored to the
cell membrane by its C-terminal
transmembrane helix, and the other folds
into two helices, and is anchored by two
fatty acid chains.
SNAP and NSF are required after fusion is
over to disassemble the SNARE
complex, so that the SNAREs can be
re-used in another round of vesicle fusion.
Synaptic vesicle fusion is blocked by
Tetanus and Botulinus toxins (the most
powerful known toxins, lethal at 0.1 ng per
kg body mass), which act by proteolytic cleavage of SNAREs. Tetatus toxin targets inhibitory neurons,
leading to spastic paralysis) whereas Botulinus toxin targets motor neurons, leading to flaccid paralysis).